Neisseria gonorrhoeae (Ng) is the causative agent of the sexually transmitted infection (STI), gonorrhea. Worldwide, >100 million new cases occur annually. Ng has become resistant to almost every antibiotic that has been used and has now achieved superbug status. The emergence and spread of ceftriaxone-resistant strains, in particular, has heralded an era of untreatable gonorrhea and represents a global public health crisis. Novel therapeutics against this pathogen are urgently needed. Complement (C2) is a critical arm of innate immune defenses against Ng. Ng evade C2 using several mechanisms, including scavenging the human C2 inhibitor, factor H (fH). fH comprises 20 domains - domains 18-20 interact with porin B and binding is augmented by lipooligosaccharide sialylation, while domains 6-7 bind to neisserial surface protein A. All Ng we have tested bind to one or both of these regions in fH. Chimeric molecules created by fusing fH domains 6-7 or 18-20 to the Fc region of IgG can effect C2-dependent killing of Ng. fH domains 18-20 in the context of full-length native fH binds to and serves to limit pathological injury to host cells, which limits the therapeutic use of 'unmodified' domains 18-20 in fH/Fc. We have overcome this limitation by introducing a point mutation in domain 19 (D1119G) of fH18-20/Fc that eliminates C2-mediated lysis of host cells without altering anti-Ng efficacy, thus providing us with a lead therapeutic molecule. In Aim 1, we will maximize the ability of the human IgG1 Fc region of fHD1119G/Fc to mediate opsonophagocytosis and C2-dependent killing of multi-drug resistant Ng isolates by introducing 3 amino acid point mutations in the Fc and eliminating the core fucose of Fc glycans. Similarly, in Aim 2a, we will optimize the effector functions of the Fc that is fused to human fH domains 6 and 7 (fH6,7/Fc) and evaluate its ability to mediate C2-dependent killing and opsonophagocytosis of drug-resistant Ng. The ability of fHD111G/Fc and fH67/Fc, which recognize distinct epitopes, to synergize and stimulate bactericidal activity against Ng will be evaluated in Aim 2b. The milestones achieved in Aims 1 and 2 (the R21 phase) will be followed by studies in vivo (R33 phase) to determine the safety, efficacy and mechanism of action of fH/Fc. Renal, ocular and hematologic toxicity (target organs for C2-mediated injury) will be assessed in Aim 3. Introducing the D1119G mutation could create a neo-epitope and elicit auto-Abs; this possibility will be addressed using our human fH transgenic (Tg) mice. In Aim 4 we will determine efficacy of the fH/Fc fusion proteins against a panel of drug-resistant Ng in the mouse model of Ng infection using our novel human fH/C4BP (C4BP; classical pathway C2 inhibitor) double Tg mice (Ng bind only human, but not mouse C2 inhibitors). Using mice that lack complement C3, C5, Fc receptors, PMNs or macrophages we will define how fH/Fc clear Ng in vivo. Many medically important microbes bind human fH through domains 6-7 and/or 18-20. While we will use Ng as a model pathogen, successful completion of these studies will provide a strong platform to assess these fH/Fc molecules against other emerging drug-resistant pathogens.